Process of Assay Selection and Optimization for the Study of Case and Control Samples from a Phase IIb Efficacy Trial of a Candidate Tuberculosis Vaccine, MVA85A
| Autor: | Harris, SA, Satti, I, Matsumiya, M, Stockdale, L, Chomka, A, Tanner, R, O'Shea, MK, Thomas, Z-R, Tameris, M, Mahomed, H, Scriba, TJ, Hanekom, WA, Fletcher, HA, McShane, H |
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| Jazyk: | angličtina |
| Rok vydání: | 2014 |
| Předmět: |
Adult
CD4-Positive T-Lymphocytes Vaccines Antigens Bacterial Enzyme-Linked Immunospot Assay Herpesvirus 4 Human Cytomegalovirus Infant Pilot Projects Mycobacterium tuberculosis CD8-Positive T-Lymphocytes Orthomyxoviridae Mycobacterium bovis Placebos Interferon-gamma South Africa Ki-67 Antigen Leukocytes Mononuclear Vaccines DNA Humans Tuberculosis Vaccines Tuberculosis Pulmonary Acyltransferases |
| Zdroj: | Clinical and Vaccine Immunology : CVI |
| ISSN: | 1556-679X 1556-6811 |
| Popis: | The first phase IIb safety and efficacy trial of a new tuberculosis vaccine since that for BCG was completed in October 2012. BCG-vaccinated South African infants were randomized to receive modified vaccinia virus Ankara, expressing the Mycobacterium tuberculosis antigen 85A (MVA85A), or placebo. MVA85A did not significantly boost the protective effect of BCG. Cryopreserved samples provide a unique opportunity for investigating the correlates of the risk of tuberculosis disease in this population. Due to the limited amount of sample available from each infant, preliminary work was necessary to determine which assays and conditions give the most useful information. Peripheral blood mononuclear cells (PBMC) were stimulated with antigen 85A (Ag85A) and purified protein derivative from M. tuberculosis in an ex vivo gamma interferon (IFN-γ) enzyme-linked immunosorbent spot assay (ELISpot) and a Ki67 proliferation assay. The effects of a 2-h or overnight rest of thawed PBMC on ELISpot responses and cell populations were determined. Both the ELISpot and Ki67 assays detected differences between the MVA85A and placebo groups, and the results correlated well. The cell numbers and ELISpot responses decreased significantly after an overnight rest, and surface flow cytometry showed a significant loss of CD4(+) and CD8(+) T cells. Of the infants tested, 50% had a positive ELISpot response to a single pool of flu, Epstein-Barr virus (EBV), and cytomegalovirus (CMV) (FEC) peptides. This pilot work has been essential in determining the assays and conditions to be used in the correlate study. Moving forward, PBMC will be rested for 2 h before assay setup. The ELISpot assay, performed in duplicate, will be selected over the Ki67 assay, and further work is needed to evaluate the effect of high FEC responses on vaccine-induced immunity and susceptibility to tuberculosis disease. |
| Databáze: | OpenAIRE |
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